CPPD crystal deposition in human articular cartilage is a common age-related event, and clinically and epidemiologically associated with degenerative joint disease, another age-related event. CPPD causes arthritis and subsequent cartilage degradation. However, the molecular mechanisms involved in CPPD formation in articular tissues remain undetermined. The goal of this grant proposal is to define the molecular basis of calcium pyrophosphate dehydrate (CPPD) crystal deposition disease. Elevated nucleotide pyrophosphohydrolase (NTPPHase) activity has been observed in CPPD-containing joint fluids and in detergent extracts of CPPD crystal-encrusted degenerated cartilage. NTPPHase catalyzes the joint fluids of PPi from ATP and other nucleotides. Several studies are consistent with the hypothesis that the bulk of PPi production occurs via the hydrolysis of extracellular ATP by ectoNTPPHase. NTPPHase exists in several molecular forms, including a 127kD sedimentable vesicle-associated NTPPHase, and two soluble forms of molecular masses of 100 and 61kD. The 127kD protein appears to represent the mature form of NTPPHase, while the 100 and 61kD forms represent soluble proteolytic fragments of the 127kD NTPPHase. The applicants have successfully cloned a partial cDNA encoding the 61kD NTPPHase fragment of the porcine 127kD NTPPHase from a porcine chondrocyte cDNA library, and also cloned its human homologue from a human chondrocyte cDNA library. Overall, their research has focused on defining the molecular properties of these NTPPHases. The Specific Aims of the current proposal are to: 1) clone the human homologue of the 127kD NTPPHase from a human chondrocyte cDNA library; 2) characterize the enzymatic activity and immunological properties of human 127kD NTPPHase; 3) molecularly characterize the gene that encodes the human NTPPHase; and 4) correlate the expression of the human NTPPHase during aging and disease. It is suggested (by the applicant) that these investigations will provide the framework for the investigators' long-term goal, which is to identify populations at risk, and to design therapeutic approaches, at either the level of transcription or inhibitors of the enzymatic activity of NTPPHase. It is hoped that these studies will lead to therapies to reduce ePPi accumulation and reduce subsequent cartilage degeneration.